南美白对虾丝氨酸蛋白酶的分离纯化及性质研究  

Purification and Characterization of Serine Proteinase from Pacific White Shrimp

作　　者：翁凌[1] 李腾[2] 阴利华[1] 孙乐常[1] 苏文金[1] 曹敏杰[1] 

机构地区：[1]集美大学生物工程学院,福建厦门361021 [2]上海海洋大学食品学院,上海201306

出　　处：《集美大学学报（自然科学版）》2010年第4期272-278,共7页Journal of Jimei University：Natural Science

基　　金：十一五国家科技支撑计划重大项目(2008BAD94B01);福建省高校水产科学技术与食品安全重点实验室基金项目(2008J401)

摘　　要：通过硫酸铵盐析、DEAE-Sepharose、Phenyl-Sepharose、Hydroxyapatite、Superdex75等方法,从南美白对虾消化腺中分离得到一种丝氨酸蛋白酶.SDS-PAGE结果显示,其分子质量约为28ku,最适pH值与最适温度分别为9.0和40℃,且pH值在7.0～10.0之间以及温度在40℃以下有较高的稳定性.底物特异性实验与抑制剂实验结果表明,该酶属于类胰蛋白酶的丝氨酸蛋白酶.动力学实验显示,以Boc-Phe-Ser-Arg-MCA为底物时,Km=0.69μmol/L,Kcat=0.33S-1,Kcat/Km=4.78×105（mol/L）-1S-1.A serine proteinase from the hepatopancreas of Pacific white shrimp was purified by a series of procedures,including ammonium sulfate precipitation,column chromatographies on DEAE-Sepharose, Phenyl-Sepharose,Hydroxyapatite and Superdex 75. Purified serine proteinase revealed a single band on SDS-PAGE. The molecular weight was about 28 ku,The optimum pH and temperature of the enzyme were 9. 0 and 40 ℃,respectivety. The enzyme was stable up to 35 ℃ and in the pH range from 7. 0 to 10. 0. Substrate specificity and inhibitor sensitivity experiments suggested that the enzyme was a trypsin-type serine proteinase. Kinetic constants of Km was 0. 69 μmol/L,Kcat was 0. 33 S-1 and Kcat /Km was 4. 78 × 105 （mol/ L）-1S-1 using Boc-Phe-Ser-Arg-MCA as substrate.

关 键 词：丝氨酸蛋白酶 南美白对虾 纯化 性质分析 

分 类 号：Q556[生物学—生物化学]